What is the significance of a negative result?
PCR. If organisms circulate only transiently during a particular phase of infection or in low numbers in peripheral blood, even
the most sensitive PCR test may not indicate that a dog is infected with a particular agent (see above "Is the organism in the sample? The clinical sensitivity of PCR" above). PCR results may also be negative because treatment has reduced the numbers of circulating organisms.
If the infecting species' DNA is not targeted by the assays' primers, PCR test results may also be negative. This can be a
problem when PCR assays target a genus. All species within a genus may not necessarily be detected by the assay.
For example, B. conradae was discovered to be a cause of canine IMHA in southern California in the early 1990s.33 However, commercial diagnostic tests targeting this species only recently became available. Since then, reemergence of this
organism as a cause of IMHA in southern California dogs has been described.7 It is important to verify with a laboratory that a species of interest for a particular patient is targeted by a given PCR
before assuming infection is ruled out based on a negative PCR test result.
Serology. Like PCR tests, serologic testing on a single sample may produce negative results in the face of infection, if the organism
causes clinical signs in the acute phase of disease, before antibodies are formed. For example, serology might not indicate
exposure in dogs acutely infected with agents such as R. rickettsii, E. canis, Babesia species, and A. phagocytophilum, because clinical signs can sometimes develop before seroconversion during infection with these agents.10,12,22,29
Furthermore, for certain organisms such Bartonella and Babesia species, antibodies may not be detectable in infected patients even after seroconversion theoretically should have occurred.7,34 The absence of antibody has been theorized to be due to young age or immunosuppression in some patients, or, for Bartonella, antigenic differences may exist between organisms grown in vitro for immunofluorescence assay (IFA) testing and organisms
infecting a patient.7,34 Importantly, up to 50% of dogs infected with Bartonella species may be seronegative.7
Sensitivity can also differ among types of serologic assays. Infrequently, in some dogs actively infected with E. canis, antibodies can be detected by using IFAs but not enzyme-linked immunosorbent assays. This may be related to differences
in the nature of antigens used in the respective assays.35
Serologic and PCR panels can facilitate the diagnosis of vector-borne disease in dogs. Clinicians should consider the epidemiology
and pathophysiology of each agent to guide testing.
Testing with both serology and PCR and testing more than one sample is often necessary to confirm the presence of infection.
Testing using both serology and PCR in parallel can improve diagnostic sensitivity and is recommended.3,4 If only one modality (PCR or serology) is used initially, and the results are negative, stored samples can be accessed to
test with an alternative methodology if both serum and EDTA samples are obtained at initial presentation. Furthermore, convalescent
serologic testing can be considered to confirm acute infection.
Repeating PCR on initial or additional samples or using specialized techniques such as enrichment before PCR testing in the
case of bartonellosis7 may help document infection in cases in which low numbers of circulating organisms is a concern.
Linda Kidd, DVM, PhD, DACVIM
College of Veterinary Medicine
Western University of Health Sciences
309 E. Second St.
Pomona, CA 91766
Shadie Ghaffari, DVM
VCA West Los Angeles Animal Hospital
1900 S. Sepulveda Blvd.
Los Angeles, CA 90025